It is presumed that the pathogenesis of peptic ulcer (mainly stomach and duodenal ulcers) results from the impairment of mucosa of the digestive tract by an imbalance between aggressive factors and defense factors. According to this presumption, even though the aggressive factors such as hydrochloric acid or pepsin are increased, the defense factors such as mucous or blood stream are also increased in the normal conditions so that mucosal tissues are not injured, thereby maintaining the homeostasis which gastric mucosa essentially possesses. When the balance is disrupted, the gastric mucosa is impaired and can result in ulcer.
Based on this mechanism, a number of anti-ulcer drugs have been developed and clinically used. For example, histamine H2 receptor blockers, which suppress gastric acid secretion, and proton pump inhibitors have been launched on the market and the results of treating ulcer have greatly progressed. Sucrose aluminum sulfate, which protects the injured mucosa and enhances the healing of ulcer though it does not inhibit gastric acid secretion has also been developed. It enjoys acceptance equivalent to that of H2 blockers. In addition, pharmaceutical compositions for treating peptic ulcer containing acid resistant fibroblast growth factor (FGF) (JP-WA-Hei 7-505736), a prophylactic and therapeutic composition for gastric mucosal disorders containing epidermal growth factor (EGF) (JP-WA-Hei 7-503471), and the like, are also under development. These growth factors were developed initially as therapeutics for treating wounds. Based on their biological activities, these factors are also being investigated for use as anti-ulcer drugs.
Recently, a new growth factor was found which is entirely different in structure from a variety of known heparin-binding growth factors. Kadomatsu et al. cloned cDNA complementary to mRNA from mouse embryonic tumor cells in which expression of mRNA was transiently enhanced at the initial stage of differentiation and induction by treatment with retinoic acid. The factor isolated was named MK 1 (Kadomatsu et al. [1988] Biochem. Biophys. Res. Commun. 151:1312-1318). The protein (hereinafter referred to as MK protein, MK polypeptide or simply MK) encoded by MK 1 cDNA is a novel protein rich in basic amino acids and cysteines and does not exhibit substantial homology to any other known proteins. Subsequently, analysis of the cDNA clone derived from mouse embryonic tumor cells indicated that there were three types of clones which had different structures at the 5' non-translational regions, but possessed an identical coding region. These three types were identified as the clones MK 1, MK 2, and MK 3. Among these clones, MK 2 was found to have the structure corresponding to the dominant mRNAs (Tomomura, M. et al. [1990] J. Biol Chem. 265:10765-10770).
Human MK cDNA has been cloned from human MK gene library using mouse MK 2 cDNA (Tsutsui, J. et al. [1991] Biochem. Biophys. Res. Commun. 176:792-797; U.S. Pat. No. 5,210,026). Sequencing revealed that human MK and mouse MK have 87% homology in their amino acid sequence.
MK protein has a molecular weight of 13 kDa and consists of 121 amino acid residues in human and 118 residues in mouse, both of which are rich in basic amino acids and cysteines. MK has some biological activities such as abilities to grow neurite, to maintain neurocytes alive, and to stimulate the fibrinolytic system in vascular endothelial cells. The MK molecule has two domains (1/2 molecule of the N-terminal and 1/2 molecule of the C-terminal domains) which are formed by the disulfide bond within the molecule. The biological activities are attributable to the C-terminal domain (Muramatsu, H. et al. [1995] Biochem. Biophys. Res. Commun. 206:468-473).
In late 1990, "heparin binding growth associated molecule" (HB-GAM), which has an amino acid sequence highly homologous to MK, was reported (Mercenmies, T. and Rauvala, H. [1990] J. Biol. Chem. 265:16721-16724). HB-GAM is also called pleiotrophin (PTN) (Li, Y. S. et al. [1990] Science 250:1690-1694), OSF-1 (Tezuka, K. et al. [1990] Biochem. Biophys. Res. Commun. 173:246-251), or heparin binding neurotrophic factor (HBNF) (Kovesde, I. et al. [1990] Biochem. Biophys. Res. Commun. 172:850-854).
HB-GAM is a protein with the molecular weight of 15 KDa and has an ability to stimulate the growth of neurites (Rauvala, H. [1989] EMBO J. 8:2933-2941). The structure of this protein is so highly preserved among species as to differ in only one amino acid between human and mouse (Li, Y. -S., et al. [1990] Science 250:1690-1694). HB-GAM has 50% homology to human MK. The positions of all cysteines are preserved in both proteins (Mercenmies, T. and Rauvala, H. [1990] J. Biol. Chem. 265:16721-16724; Li, Y. -S., et al. [1990] Science 250:1690-1694) as is the disulfide bond (Fabri, L. et al. [1992] Biochem. Int. 28:1-9). Particularly, MK and HB-GAM are highly homologous in the two domains surrounding the S-S bond. These results indicate that MK and HB-GAM form a new family as heparin binding growth factors (Muramatsu, T. et al. [1994] Dev. Growth Differ. 36:1-8).
It has also been reported that the expression level of MK increased in a variety of human cancers (Tsutsui, J. et al. [1993] Cancer Res. 53:1281-1285). In all six cases of Wilms' tumor, MK was found not to be expressed. In the patients with liver or esophagus cancer, MK was not expressed in noncancerous areas but was often highly expressed in cancerous areas. In cases of colon and stomach cancers, the expression of MK was observed often even in the noncancerous areas, but strong expression mostly occurred in the cancerous areas. MK was also expressed in lung cancer, breast cancer, and neuroblastoma. Thus, the increase in the MK expression level is considered to correlate with the progress of cancer.
It was also reported that, in experimental cerebral infarction in rats, MK was expressed around the area of infarction and also appeared in the edematic area that recovered later without proceeding to necrosis at such an early stage as one day after the onset of infarction (Yoshida, Y. et al. [1995] Dev. Brain Res. 85:25-30). These results suggest that MK plays a very important role not only in the process of embryonic development but also in the repair of tissues. A disorder of the retina caused by continuous light irradiation was found to be ameliorated by the injection of MK (Unoki, K. et al. [1994] Inv. Ophthal. Vis. Sci. 35:4603-4068). Furthermore, in the brain of patients with Alzheimer's disease, MK was detected in the senile plaques without exception (Yasuhara, O. et al. [1993] Biochem. Biophys. Res. Commun. 192:246-251). A recent report demonstrated that, as a result of treating cultured normal human skin fibroblasts with a variety of concentrations of MK, a treatment at 60 ng/ml for 72 hours enhanced synthesis of collagen and mucopolysaccharide to twice the normal level (Tajima, S. et al. [1995] Seikagaku 67(7):938; Yamada, H. et al. [1997] Arch. Dermatol. Res. 289:429-433).
There are many anti-ulcer drugs which inhibit acid secretion or increase defense factors, but no drug has been developed yet which can achieve an ultimate object of anti-ulcer drugs, which is to completely cure ulcers without causing recurrence.
It can be said that currently used H2 blockers and proton pump inhibitors are, in terms of a cure, dependent upon autotherapy due to a natural increase of the cells. Under such circumstances, the growth factors such as EGF and FGF have been launched to aim at curing the ulcer nearly to the previous level by enhancing the growth of cells at ulcerous regions.
The in vivo mechanism of these factors for the healing of ulcer is as follows. First, fibroblasts proliferate by the action of EGF, FGF, etc. at ulcerous regions with the simultaneous occurrence of angiogenesis by the action of FGF and the like, followed by granulation. Then, TGF-.alpha., EGF, etc. cause migration and growth of epithelial cells, accompanied by disappearance of granulation. Thus, the ulcerous regions recover to the normal conditions.
Since this healing mechanism usually involves substances that the living body inherently possesses, no particular problem is expected to occur. However, when EGF and FGF are forcibly given, there is a possibility that undesirable effects take place. For example, when EGF is administered, only epithelial cells are healed without enough granulation. In the case of administration of FGF, excessive granulation may possibly be induced, thereby failing to heal to the normal state but possibly causing recurrence of ulcer.